Flu mutations in a single cell help to predict immune response

Influenza A virus has a broad cellular tropism in the respiratory tract. Infected epithelial cells sense the infection and initiate an antiviral response. Here, we used single-cycle replication reporter viruses to analyze the early cellular response to influenza infection in vivo. This approach revealed distinct tiers of antiviral responses that were associated with the magnitude of virus replication. We also unveiled disparate protection of epithelial cell types mediated by interferon during virus spread. These results demonstrate the early landscape of virus–host interactions in vivo with the magnitude and round of replication revealing distinct antiviral signatures and responses. Influenza virus has a broad cellular tropism in the respiratory tract. Infected epithelial cells sense the infection and initiate an antiviral response. To define the antiviral response at the earliest stages of infection we used a series of single-cycle reporter viruses. These viral probes demonstrated cells in vivo harbor a range in magnitude of virus replication. Transcriptional profiling of cells supporting different levels of replication revealed tiers of IFN-stimulated gene expression. Uninfected cells and cells with blunted replication expressed a distinct and potentially protective antiviral signature, while cells with high replication expressed a unique reserve set of antiviral genes. Finally, we used these single-cycle reporter viruses to determine the antiviral landscape during virus spread, which unveiled disparate protection of epithelial cell subsets mediated by IFN in vivo. Together these results highlight the complexity of virus–host interactions within the infected lung and suggest that magnitude and round of replication tune the antiviral response.

The outcome of viral infection is extremely heterogeneous, with infected cells only sometimes activating innate immunity. Here we assess how the genetic variation inherent in viral populations contributes to this heterogeneity. We do this by developing a new approach to determine both the transcriptome and full-length sequences of all viral genes in single influenza-infected cells. Infections that activate an innate-immune response in single cells are associated with viral defects that include amino-acid mutations, internal deletions, and failure to express key genes. However, immune activation remains stochastic in cells infected by virions with these defects, and sometimes occurs even in cells infected by virions that express unmutated copies of all genes. Our work shows that the genetic variation present in influenza virus populations substantially contributes to but does not fully explain the heterogeneity in infection outcome and immune activation in single infected cells.